Reduced Representation Genome Sequencing Research Articles

Discovery of a new gall-inducing species, Aciurina luminaria (Insecta, Diptera, Tephritidae) via multi-trait integrative taxonomy

Integrative taxonomic practices that combine multiple lines of evidence for species delimitation greatly improve our understanding of intra- and inter-species variation and biodiversity. However, extended phenotypes remain underutilized despite their potential as a species-specific set of extracorporeal morphological and life history traits. Primarily relying on variations in wing patterns has caused taxonomic confusion in the genus Aciurina, which are gall-inducing flies on Asteraceae plants in western North America. However, species display distinct gall morphologies that can be crucial for species identification. Here we investigate a unique gall morphotype in New Mexico and Colorado that was previously described as a variant of that induced by Aciurina bigeloviae (Cockerell, 1890). Our analysis has discovered several consistent features that distinguish it from galls of A. bigeloviae. A comprehensive description of Aciurina luminaria Baine, sp. nov. and its gall is provided through integrative taxonomic study of gall morphology, host plant ecology, wing morphometrics, and reduced-representation genome sequencing.

Analysis of Elimination Effects of Inbreeding on Genotype Frequency in Larval Stages of Chinese Shrimp.

Marine animals possess genomes of considerable complexity and heterozygosity. Their unique reproductive system, characterized by high fecundity and substantial early mortality rates, increases the risk of inbreeding, potentially leading to severe inbreeding depression during various larval developmental stages. In this study, we established a set of inbred families of Fenneropenaeus chinensis, with an inbreeding coefficient of 0.25, and investigated elimination patterns and the manifestations of inbreeding depression during major larval developmental stages. Reduced-representation genome sequencing was utilized to explore the genotype frequency characteristics across two typical elimination stages. The results revealed notable mortality in hatching and metamorphosis into mysis and post-larvae stages. Inbreeding depression was also evident during these developmental stages, with depression rates of 24.36%, 29.23%, and 45.28%. Segregation analysis of SNPs indicated an important role of gametic selection before hatching, accounting for 45.95% of deviation in the zoea stage. During the zygotic selection phase of larval development, homozygote deficiency and heterozygote excess were the main selection types. Summation of the two types explained 82.31% and 89.91% of zygotic selection in the mysis and post-larvae stage, respectively. The overall distortion ratio decreased from 22.37% to 12.86% in the late developmental stage. A total of 783 loci were identified through selective sweep analysis. We also found the types of distortion at the same locus could change after the post-larvae stage. The predominant shifts included a transition of gametic selection toward normal segregation and other forms of distortion to heterozygous excess. This may be attributed to high-intensity selection on deleterious alleles and genetic hitchhiking effects. Following larval elimination, a greater proportion of heterozygous individuals were preserved. We detected an increase in genetic diversity parameters such as expected heterozygosity, observed heterozygosity, and polymorphic information content in the post-larvae stage. These findings suggest the presence of numerous recessive deleterious alleles and their linkage and suggest a major role of the partial dominance hypothesis. The results provide valuable insights into the mechanisms of inbreeding depression in marine animals and offer guidance for formulating breeding strategies in shrimp populations.

Genomic analysis of wolves from Pakistan clarifies boundaries among three divergent wolf lineages.

Among the three main divergent lineages of gray wolf (Canis lupus), the Holarctic lineage is the most widespread and best studied, particularly in North America and Europe. Less is known about Tibetan (also called Himalayan) and Indian wolf lineages in southern Asia, especially in areas surrounding Pakistan where all three lineages are thought to meet. Given the endangered status of the Indian wolf in neighboring India and unclear southwestern boundary of the Tibetan wolf range, we conducted mitochondrial and genome-wide sequencing of wolves from Pakistan and Kyrgyzstan. Sequences of the mitochondrial D-loop region of 81 wolves from Pakistan indicated contact zones between Holarctic and Indian lineages across the northern and western mountains of Pakistan. Reduced-representation genome sequencing of eight wolves indicated an east-to-west cline of Indian to Holarctic ancestry, consistent with a contact zone between these two lineages in Pakistan. The western boundary of the Tibetan lineage corresponded to the Ladakh region of India's Himalayas with a narrow zone of admixture spanning this boundary from the Karakoram Mountains of northern Pakistan into Ladakh, India. Our results highlight the conservation significance of Pakistan's wolf populations, especially the remaining populations in Sindh and Southern Punjab that represent the highly endangered Indian lineage.

Global phylogenomic assessment of Leptoseris and Agaricia reveals substantial undescribed diversity at mesophotic depths

BackgroundMesophotic coral communities are increasingly gaining attention for the unique biological diversity they host, exemplified by the numerous mesophotic fish species that continue to be discovered. In contrast, many of the photosynthetic scleractinian corals observed at mesophotic depths are assumed to be depth-generalists, with very few species characterised as mesophotic-specialists. This presumed lack of a specialised community remains largely untested, as phylogenetic studies on corals have rarely included mesophotic samples and have long suffered from resolution issues associated with traditional sequence markers.ResultsHere, we used reduced-representation genome sequencing to conduct a phylogenomic assessment of the two dominant mesophotic genera of plating corals in the Indo-Pacific and Western Atlantic, respectively, Leptoseris and Agaricia. While these genome-wide phylogenies broadly corroborated the morphological taxonomy, they also exposed deep divergences within the two genera and undescribed diversity across the current taxonomic species. Five of the eight focal species consisted of at least two sympatric and genetically distinct lineages, which were consistently detected across different methods.ConclusionsThe repeated observation of genetically divergent lineages associated with mesophotic depths highlights that there may be many more mesophotic-specialist coral species than currently acknowledged and that an urgent assessment of this largely unstudied biological diversity is warranted.

Domestication and Genetic Diversity Analysis of Several Sheep Breeds in China

Sheep is the main livestock species in northern China. Under diverse climatic conditions, a gene pool of local sheep resources is now forming. There are many sheep breeds in northern China, including local breeds and hybrid breeds. However, little is known about the genetic background and genetic variation of local sheep populations in northern China. Therefore, our aims is to provide a characterization of the population structure and genetic relationship within sheep breeds in northern China, and to study its genetic relationship with other sheep breeds. We collected blood samples from 390 sheep for genetic diversity analysis using Reduced-Representation Genome Sequencing (RRGS). This study used bioinformatics methods to analyze linkage disequilibrium, population structure, evolutionary tree, selective sweep etc. The mixed model revealed in this study may be formed in the process of sheep breeding, partially overlapping with historical and archaeological findings. New information on the domestication and genetic variety of sheep is provided by this study, which will help sheep in the future be improved through genomics.

Phylogeny of the Diploid Species of Rubus (Rosaceae).

Rubus L. (Rosaceae, Rosoideae) contains around 700 species distributed on all continents except Antarctica, with the highest species diversity in temperate to subtropical regions of the northern hemisphere. The taxonomy of Rubus is challenging due to the frequency of polyploidy, hybridization and apomixis. Previous studies mostly sampled sparsely and used limited DNA sequence data. The evolutionary relationships between infrageneric taxa, therefore, remain to be further clarified. In the present study, genotyping by sequencing (GBS) reduced-representation genome sequencing data from 186 accessions representing 65 species, 1 subspecies and 17 varieties of Rubus, with emphasis on diploid species, were used to infer a phylogeny using maximum likelihood and maximum parsimony methods. The major results were as follows: (1) we confirmed or reconfirmed the polyphyly or paraphyly of some traditionally circumscribed subgenera, sections and subsections; (2) 19 well-supported clades, which differed from one another on molecular, morphological and geographical grounds, were identified for the species sampled; (3) characteristics such as plants with dense bristles or not, leaves leathery or papyraceous, number of carpels, instead of inflorescences paniculate or not, aggregate fruits and leaves abaxially tomentose or not, may be of some use in classifying taxa whose drupelets are united into a thimble-shaped aggregate fruit that falls in its entirety from the dry receptacle; and (4) a preliminary classification scheme of diploid species of Rubus is proposed based on our results combined with those from previous phylogenetic analyses.

Community-wide correlations between species richness, abundance and population genomic diversity in a freshwater biodiversity hotspot.

Understanding patterns of diversity across macro (e.g. species-level) and micro (e.g. molecular-level) scales can shed light on community function and stability by elucidating the abiotic and biotic drivers of diversity within ecological communities. We examined the relationships among taxonomic and genetic metrics of diversity in freshwater mussels (Bivalvia: Unionidae), an ecologically important and species-rich group in the southeastern United States. Using quantitative community surveys and reduced-representation genome sequencing across 22 sites in seven rivers and two river basins, we surveyed 68 mussel species and sequenced 23 of these species to characterize intrapopulation genetic variation. We tested for the presence of species diversity-abundance correlations (i.e. the more-individuals hypothesis, MIH), species-genetic diversity correlations (SGDCs) and abundance-genetic diversity correlations (AGDCs) across all sites to evaluate relationships between different metrics of diversity. Sites with greater cumulative multispecies density (a standardized metric of abundance) had a greater number of species, consistent with the MIH hypothesis. Intrapopulation genetic diversity was strongly associated with the density of most species, indicating the presence of AGDCs. However, there was no consistent evidence for SGDCs. Although sites with greater overall densities of mussels had greater species richness, sites with higher genetic diversity did not always exhibit positive correlations with species richness, suggesting that there are spatial and evolutionary scales at which the processes influencing community-level diversity and intraspecific diversity differ. Our work reveals the importance of local abundance as indicator (and possibly a driver) of intrapopulation genetic diversity.

Triploid and aneuploid hybrids obtained from hybridization between Eriobotrya japonica and E. cavaleriei

A short ripening stage, multiple large seeds per fruit, and anthracnose disease seriously restrict the loquat (Eriobotrya japonica) industry. To obtain breeding germplasm of loquat with multiple ideal traits, a wild Eriobotrya species, E. cavaleriei (2n=2x=34), which flowers in late spring and early summer and whose fruits ripen in autumn, was used as the male parent to produce triploid plants from crosses with loquat tetraploid (2n=4x=68) lines (EjK474, EjB433 and EjB456). Fifteen progenies were obtained from three hybridizations, and 13 plants were identified as hybrids according to insertion-deletion (InDel) markers. All 13 hybrids were roughly identified as triploids by flow cytometry. Therein, 4 plants were identified as aneuploids with decreased or increased chromosome numbers based on sequencing depth according to reduced-representation genome sequencing. The sequencing depth on some chromosomes of these 4 hybrids was approximately 2/3 times or 1.41 times the average depth of all detected InDel loci. Frequencies of the loquat parent-specific alleles on increased or decreased chromosomes were approximately 50% or 75%. This verified that increased or decreased alleles in the 4 hybrids were all from tetraploid parents. The fruit flesh color genotypes of all hybrids were detected; there were two red-flesh alleles and one white-flesh allele in all hybrids. Leaves of E. cavaleriei were highly resistant to anthracnose in an in vitro assay, and the leaves of loquat B433 were sensitive. Disease spot diameters on the leaves of hybrids were significantly smaller (p < 0.01) than those of B433 after infection by spores of C. gloeosporioides. Therefore, the resistance of the hybrids was greater than that of the female parent B433. All these results showed that allotriploids can be obtained via hybridization between loquat tetraploids and E. cavaleriei, with some aneuploids as secondary products. Different flowering phases and fruit ripening stages, seedlessness and resistance to anthracnose might be combined in allotriploids. This study verified that resequencing can be used to identify the molecular karyotype of interspecific hybrids.

Using landscape genomics to assess local adaptation and genomic vulnerability of a perennial herb Tetrastigma hemsleyanum (Vitaceae) in subtropical China.

Understanding adaptive genetic variation of plant populations and their vulnerabilities to climate change are critical to preserve biodiversity and subsequent management interventions. To this end, landscape genomics may represent a cost-efficient approach for investigating molecular signatures underlying local adaptation. Tetrastigma hemsleyanum is, in its native habitat, a widespread perennial herb of warm-temperate evergreen forest in subtropical China. Its ecological and medicinal values constitute a significant revenue for local human populations and ecosystem. Using 30,252 single nucleotide polymorphisms (SNPs) derived from reduced-representation genome sequencing in 156 samples from 24 sites, we conducted a landscape genomics study of the T. hemsleyanum to elucidate its genomic variation across multiple climate gradients and genomic vulnerability to future climate change. Multivariate methods identified that climatic variation explained more genomic variation than that of geographical distance, which implied that local adaptation to heterogeneous environment might represent an important source of genomic variation. Among these climate variables, winter precipitation was the strongest predictor of the contemporary genetic structure. F ST outlier tests and environment association analysis totally identified 275 candidate adaptive SNPs along the genetic and environmental gradients. SNP annotations of these putatively adaptive loci uncovered gene functions associated with modulating flowering time and regulating plant response to abiotic stresses, which have implications for breeding and other special agricultural aims on the basis of these selection signatures. Critically, modelling revealed that the high genomic vulnerability of our focal species via a mismatch between current and future genotype-environment relationships located in central-northern region of the T. hemsleyanum's range, where populations require proactive management efforts such as assistant adaptation to cope with ongoing climate change. Taken together, our results provide robust evidence of local climate adaption for T. hemsleyanum and further deepen our understanding of adaptation basis of herbs in subtropical China.

Strategies for Intraspecific Genotyping of Duckweed: Comparison of Five Orthogonal Methods Applied to the Giant Duckweed Spirodela polyrhiza.

The predominantly vegetative propagating duckweeds are of growing commercial interest. Since clonal accessions within a respective species can vary considerably with respect to their physiological as well as biochemical traits, it is critical to be able to track the clones of species of interest after their characterization. Here, we compared the efficacy of five different genotyping methods for Spirodela polyrhiza, a species with very low intraspecific sequence variations, including polymorphic NB-ARC-related loci, tubulin-gene-based polymorphism (TBP), simple sequence repeat variations (SSR), multiplexed ISSR genotyping by sequencing (MIG-seq), and low-coverage, reduced-representation genome sequencing (GBS). Four of the five approaches could distinguish 20 to 22 genotypes out of the 23 investigated clones, while TBP resolved just seven genotypes. The choice for a particular method for intraspecific genotyping can depend on the research question and the project budget, while the combination of orthogonal methods may increase the confidence and resolution for the results obtained.

Hybridization between closely related songbirds is related to human habitat disturbance.

Human habitat disturbances can promote hybridization between closely related, but typically reproductively isolated, species. We explored whether human habitat disturbances are related to hybridization between two closely related songbirds, black-capped and mountain chickadees, using both genomic and citizen science data sets. First, we genotyped 409 individuals from across both species' ranges using reduced-representation genome sequencing and compared measures of genetic admixture to a composite measure of human landscape disturbance. Then, using eBird observations, we compared human landscape disturbance values for sites where phenotypically diagnosed hybrids were observed to locations where either parental species was observed to determine whether hybrid chickadees are reported in more disturbed areas. We found that hybridization between black-capped and mountain chickadees positively correlates with human habitat disturbances. From genomic data, we found that (1) hybrid index (HI) significantly increased with habitat disturbance, (2) more hybrids were sampled in disturbed habitats, (3) mean HIs were higher in disturbed habitats versus wild habitats, and (4) hybrids were detected in habitats with significantly higher disturbance values than parentals. Using eBird data, we found that both hybrid and black-capped chickadees were significantly more disturbance-associated than mountain chickadees. Surprisingly, we found that nearly every black-capped chickadee we sampled contained some proportion of hybrid ancestry, while we detected very few mountain chickadee backcrosses. Our results highlight that hybridization between black-capped and mountain chickadees is widespread, but initial hybridization is rare (few F1s were detected). We conclude that human habitat disturbances can erode pre-zygotic reproductive barriers between chickadees and that post-zygotic isolation is incomplete. Understanding what becomes of recently hybridizing species following large-scale habitat disturbances is a new, but pressing, consideration for successfully preserving genetic biodiversity in a rapidly changing world.

Identification of Signatures of Selection for Litter Size and Pubertal Initiation in Two Sheep Populations.

Simple SummaryFecundity is an important economic trait of sheep that directly affects their economic and productive efficiency. Our study aimed to identify single nucleotide polymorphism (SNP) loci associated with sheep puberty or litter size which could be used in future breeding programs to improve fertility. We obtained one SNP on GNAQ gene (NC_040253.1: g.62677376G > A), which was associated with litter size in Cele Black sheep. Perhaps this SNP could be a criterion to screen polytocous sheep.Fecundity is an important economic trait in sheep that directly affects their economic and productive efficiency. Our study aimed to identify SNP loci associated with sheep puberty or litter size which could be used in future breeding programs to improve fertility. Genomic DNA was obtained from Hetian and Cele Black sheep breeds and used for reduced-representation genome sequencing to identify SNP loci associated with pubertal initiation and litter size. Selective signatures analysis was performed based on the fixation index and nucleotide diversity, followed by pathway analysis of the genes contained in the selected regions. The selected SNP loci in the genes associated with pubertal initiation and litter size were validated using both sheep breeds. In total, 384,718 high quality SNPs were obtained and 376 genes were selected. Functional annotation of genes and enrichment analysis identified 12 genes associated with pubertal initiation and 11 genes associated with litter size. SNP locus validation showed that two SNP on PAK1 and four on ADCY1 may be associated with pubertal initiation, and one SNP on GNAQ gene (NC_040253.1: g.62677376G > A) was associated with litter size in Cele Black sheep. Our results provide new theoretical support for sheep breeding.

A novel multifunctional haplotyping-based preimplantation genetic testing for different genetic conditions.

Is there an efficient and cost-effective detection platform for different genetic conditions about embryos? A multifunctional haplotyping-based preimplantation genetic testing platform was provided for detecting different genetic conditions. Genetic disease and chromosomal rearrangement have been known to significantly impact fertility and development. Therefore, preimplantation genetic testing for aneuploidy (PGT-A), monogenic disorders (PGT-M) and structural rearrangements (PGT-SR), a part of ART, has been presented together to minimize the fetal genetic risk and increase pregnancy rate. For patients or their families who are suffering from chromosome abnormality, monogenic disease, unexplained repeated spontaneous abortion or implantation failure, after accepting genetic counseling, they may be suggested to accept detection from more than one PGT platforms about the embryos to avoid some genetic diseases. However, PGT platforms work through different workflows. The high costliness, lack of material and long-time operation of combined PGT platforms limit their application. All 188 embryonic samples from 43 families were tested with HaploPGT platform, and most of their genetic abnormalities had been determined by different conventional PGT methods beforehand. Among them, there were 12 families only carrying structural rearrangements (115 embryos) in which 9 families accepted implantation and 5 families had normal labor ART outcomes, 7 families only carrying monogenic diseases (26 embryos) and 3 families carrying both structural rearrangements and monogenic diseases (26 embryos). Twelve monopronucleated zygotes (1PN) samples and 9 suspected triploid samples were collected from 21 families. Here, we raised a comprehensive PGT method called HaploPGT, combining reduced representation genome sequencing, read-count analysis, B allele frequency and haplotyping analysis, to simultaneously detect different genetic disorders in one single test. With 80 million reads (80M) genomic data, the proportion of windows (1 million base pairs (Mb)) containing two or more informative single nucleotide polymorphism (SNP) sites was 97.81%, meanwhile the genotyping error rate stabilized at a low level (2.19%). Furthermore, the informative SNPs were equally distributed across the genome, and whole-genomic haplotyping was established. Therefore, 80M was chosen to balance the cost and accuracy in HaploPGT. HaploPGT was able to identify abnormal embryos with triploid, global and partial loss of heterozygosity, and even to distinguish parental origin of copy number variation in mosaic and non-mosaic embryos. Besides, by retrospectively analyzing 188 embryonic samples from 43 families, HaploPGT revealed 100% concordance with the available results obtained from reference methods, including PGT-A, PGT-M, PGT-SR and PGT-HLA. Despite the numerous benefits HaploPGT could bring, it still required additional family members to deduce the parental haplotype for identifying balanced translocation and monogenic mutation in tested embryos. In terms of PGT-SR, the additional family member could be a reference embryo with unbalanced translocation. For PGT-M, a proband was normally required. In both cases, genomic information from grandparents or parental siblings might help for haplotyping theoretically. Another restriction was that haploid, and diploid resulting from the duplication of a haploid, could not be told apart by HaploPGT, but it was able to recognize partial loss of heterozygosity in the embryonic genome. In addition, it should be noted that the location of rearrangement breakpoints and the situation of mutation sites were complicated, which meant that partial genetic disorders might not be completely detected. HaploPGT is an efficient and cost-effective detection platform with high clinical value for detecting genetic status. This platform could promote the application of PGT in ART, to increase pregnancy rate and decrease the birth of children with genetic diseases. This study was supported by grants from the National Natural Science Foundation of China (81873478, to L.H.), National Key R&D Program of China (2018YFC1003100, to L.H.), the Natural Science Foundation of Hunan Province (Grant 2022JJ30414, to P.X.), Hunan Provincial Grant for Innovative Province Construction (2019SK4012) and the Scientific Research Foundation of Reproductive and Genetic Hospital of China International Trust & Investment Corporation (CITIC)-Xiangya (YNXM-201910). Haplotyping analysis has been licensed to Basecare Co., Ltd. L.K., Y.M., K.K., D.Z., N.L., J.Z. and R.D. are Basecare Co., Ltd employees. The other authors declare no competing interests. N/A.

Phylogenetic Reconstruction of the Rainforest Lineage Fontainea Heckel (Euphorbiaceae) Based on Chloroplast DNA Sequences and Reduced-Representation SNP Markers

Fontainea is a plant genus with nine recognised species that occur across the tropical and subtropical rainforests of Australia, Papua New Guinea, New Caledonia, and Vanuatu. One of these species is cultivated commercially as the source of a cancer therapeutic, and several other species are under threat of extinction. Despite this, the phylogenetic relationships of the genus have not been explored. Our study assessed the phylogeny of seven Fontainea taxa from the Australian and Pacific Island complex using chloroplast DNA sequence data and reduced-representation genome sequencing. Maximum-likelihood and consensus network trees were used to infer the topology of phylogenetic relationships between species, which highlighted three distinct lineages and a number of sister species. Our results indicated that the geographically disjunct species Fontainea venosa and F. pancheri formed a sister group at the earliest position of divergence for the genus. The data also revealed that the vulnerable Fontainea australis and the critically endangered F. oraria form a sister subclade with evidence of some shared plastid genotypes. Generally, our phylogenetic reconstruction supports the modern taxonomical nomenclature. However, we suggest further accessions across several species may support improved genetic distinctions between the sister groups of Fontainea within the genus.

The tangled evolutionary history of a long-debated Mesoamerican taxon: The Velazquez Woodpecker (Melanerpes santacruzi, Aves: Picidae)

The Velazquez Woodpecker Melanerpes santacruzi is a highly polytypic species distributed from east-central Mexico to northern Nicaragua. The ample variation in body size, barring of the plumage, and the coloration of nasal tufts, neck, and belly have fueled debates about the taxonomy and evolutionary history of the species; however, the processes generating these patterns of variation and the underlying population dynamics throughout the species’ distribution remain poorly understood. Here, we employed reduced representation genome sequencing (NextRAD) and Ecological Niche Modeling methods to test the distinctiveness of the Velazquez Woodpecker based on this new set of genomic data and analyze the correspondence of the genetic structure and ecological differentiation with phenotypic variation and geographic distribution. From phylogenetic and demographic analyses including the Golden-Fronted (M. aurifrons) and Red-bellied Woodpecker (M. carolinus), we obtained results congruent with previous molecular phylogenies. The clades of M. santacruzi and M. carolinus-M. aurifrons are reciprocally monophyletic, although the sister group relationship of M. aurifrons is ambiguous. Using genetic and ecological analyses, we found that the species is structured into three genetically and ecologically differentiated groups comprising the subspecies (1) M. s. santacruzi, (2) M. s. dubius and (3) M. s. grateloupensis-polygrammus-veraecrucis. These groups diverged recently, with two splits between 250,000 and 150,000 years ago, and show a significant genetic admixture among them, especially in their current contact zones. Ecological and demographic models suggest the existence of intermittent areas of sympatry and connectivity among populations of M. santacruzi since the Last Interglacial period. We also found evidence of bi-directional gene flow between the species M. aurifrons and the nearby populations of M. santacruzi (M. s. grateloupensis), along the Sierra Madre Oriental in northeastern Mexico. Gene flow seems to be uneven, with prevalence of movement in the direction from M. aurifrons to M. s. grateloupensis.

Identification of novel Z/W chromosome-specific markers from the giant freshwater prawn Macrobrachium rosenbergii

The giant freshwater prawn (Macrobrachium rosenbergii) is a commercially cultured species in China. A sex chromosome-specific marker is crucial for sex identification and distinguishing sex chromosomes. Here, based on the results of reduced-representation genome sequencing and genome survey of M. rosenbergii, we identified steady sex-linked markers. Further nucleotide alignment analysis revealed 83.42% nucleotide similarity between the amplified W- and Z-chromosome fragments. qPCR confirmed that Z chromosome-specific fragments were present in males approximately twice as often compared to females. Similarly, the W chromosome-specific fragments were present in super females approximately twice as often compared to females. Our findings elucidate the ZW/ZZ sex determination type and contribute to exploring sex-related or sex-determining genes present on the sex chromosomes of M. rosenbergii.

Application of Restriction Site-Associated DNA Sequencing (RAD-Seq) for Copy Number Variation and Triploidy Detection in Human

At present, low-pass whole-genome sequencing (WGS) is frequently used in clinical research and in the screening of copy number variations (CNVs). However, there are still some challenges in the detection of triploids. Restriction site-associated DNA sequencing (RAD-Seq) technology is a reduced-representation genome sequencing technology developed based on next-generation sequencing. Here, we verified whether RAD-Seq could be employed to detect CNVs and triploids. In this study, genomic DNA of 11 samples was extracted employing a routine method and used to build libraries. Five cell lines of known karyotypes and 6 triploid abortion tissue samples were included for RAD-Seq testing. The triploid samples were confirmed by STR analysis and also tested by low-pass WGS. The accuracy and efficiency of detecting CNVs and triploids by RAD-Seq were then assessed, compared with low-pass WGS. In our results, RAD-Seq detected 11 out of 11 (100%) chromosomal abnormalities, including 4 deletions and 1 aneuploidy in the purchased cell lines and all triploid samples. By contrast, these triploids were missed by low-pass WGS. Furthermore, RAD-Seq showed a higher resolution and more accurate allele frequency in the detection of triploids than low-pass WGS. Our study shows that, compared with low-pass WGS, RAD-Seq has relatively higher accuracy in CNV detection at a similar cost and is capable of identifying triploids. Therefore, the application of this technique in medical genetics has a significant potential value.

Generation of unequal nuclear genotype proportions in Rhizophagus irregularis progeny causes allelic imbalance in gene transcription.

Summary Arbuscular mycorrhizal fungi (AMF) form mutualisms with most plant species. The model AMF Rhizophagus irregularis is common in many ecosystems and naturally forms homokaryons and dikaryons. Quantitative variation in allele frequencies in clonally dikaryon offspring suggests they disproportionately inherit two distinct nuclear genotypes from their parent. This is interesting, because such progeny strongly and differentially affect plant growth. Neither the frequency and magnitude of this occurrence nor its effect on gene transcription are known.Using reduced representation genome sequencing, transcriptomics, and quantitative analysis tools, we show that progeny of homokaryons and dikaryons are qualitatively genetically identical to the parent. However, dikaryon progeny differ quantitatively due to unequal inheritance of nuclear genotypes. Allele frequencies of actively transcribed biallelic genes resembled the frequencies of the two nuclear genotypes.More biallelic genes showed transcription of both alleles than monoallelic transcription, but biallelic transcription was less likely with greater allelic divergence. Monoallelic transcription levels of biallelic genes were reduced compared with biallelic gene transcription, a finding consistent with genomic conflict.Given that genetic variation in R. irregularis is associated with plant growth, our results establish quantitative genetic variation as a future consideration when selecting AMF lines to improve plant production.

Population genomic analyses of schistosome parasites highlight critical challenges facing endgame elimination efforts

Schistosomiasis persists in Asian regions despite aggressive elimination measures. To identify factors enabling continued parasite transmission, we performed reduced representation genome sequencing on Schistosoma japonicum miracidia collected across multiple years from transmission hotspots in Sichuan, China. We discovered strong geographic structure, suggesting that local, rather than imported, reservoirs are key sources of persistent infections in the region. At the village level, parasites collected after referral for praziquantel treatment are closely related to local pre-treatment populations. Schistosomes within villages are also highly related, suggesting that only a few parasites from a limited number of hosts drive re-infection. The close familial relationships among miracidia from different human hosts also implicate short transmission routes among humans. At the individual host level, genetic evidence indicates that multiple humans retained infections following referral for treatment. Our findings suggest that end-game schistosomiasis control measures should focus on completely extirpating local parasite reservoirs and confirming successful treatment of infected human hosts.

Sex assignment in a non-model organism in the absence of field records using Diversity Arrays Technology (DArT) data

Conservation genomics research often relies on accurate sex information to make inferences about species demography, dispersal, and population structure. However, field determined sex data are not always available and can be subject to human error, while laboratory sex assignment methods such as PCR assays can often be costly and challenging for non-model species. Conservation genomics programs increasingly use reduced-representation genome sequencing to assess neutral and functional genetic diversity, population structure, gene flow and pedigrees in threatened species. Here we demonstrate that sex can be determined from reduced-representation sequencing data produced by the increasingly popular Diversity Arrays Technology sequencing workflow (DArT-seq) using a program originally designed for application to shotgun data. This program—sexassign—compares the “dosage” of sequencing reads mapping to autosomes versus the X chromosome. In the present study, sexassign was used to identify the sex of 60 field-collected Greater Stick-Nest Rat (Leporillus conditor) samples, despite the absence of an annotated reference genome for the species. This “read-dosage” approach is not only more accurate and affordable than traditional sex assignment methods, but can be applied to any diploid organism with a heterogametic sex determination system—including non-model and understudied species of conservation importance—by using FASTQs generated by DArT.